The study of the mechanisms whereby normal and tumor cells coordinate growth with nutrient availability is a fundamental problem in cell biology. Cells need to adapt and respond to changes in nutrient status to maintain cell growth and metabolic homeostasis, and to cope with stress. Deregulation of the cellular response to nutrients impacts tumorigenesis and can be a novel source of therapeutics in cancer. mTORC1 is a central kinase in nutrient sensing by regulating fundamental processes in cancer cell progression such as growth and autophagy. Multiple studies have addressed the mechanisms of mTORC1 activation by growth factors, insulin and energy levels. However, the signaling mechanisms mediating the amino acids response are not well understood. This grant application is based on our recently published findings demonstrating that the signaling adapter and autophagy substrate, p62, is a novel critical modulator of mTORC1 activation by amino acids. This is of great significance because by activating this pathway p62 regulates cell size and growth, and emerges as a key node in nutrient sensing and autophagy modulation, all essential events in normal homeostasis and tumorigenesis. However, the precise mechanisms of mTORC1 regulation by nutrients and the role of p62 in this process still remain to be determined. In this regard, our preliminary data have identified MEKK3 as a novel partner of p62 for mTORC1 in response to amino acids, as well as that p62 is phosphorylated by a novel amino acids-stimulated MEKK3-dependent cascade, which is critical for its activation. The long-term goal of this proposal is to understand, at a molecular and cellular level, the mechanisms whereby cells translate nutrient-sensing signals, specifically those of amino acids, for the activation of mTORC1, a critical event in cancer progression. The specific goal of this application is to rigorously test the hypothesis that the new p62/MEKK3 complex, together with the ability of p62 to get phosphorylated in response to nutrients, is a central process in cancer through the activation of mTORC1. These goals will be addressed in three specific aims: 1) Determine the role and mechanisms of action of the PB1-containing MEKK3 in p62-regulated mTORC1 activation; 2) Determine the role of p62 phosphorylation as a nutrient sensor; and 3) Establish the functional role of the novel p62-MEKK3 nutrient sensing pathway in cancer. The results of all these studies will provide significantly impactful conceptual advance to our understanding of how cells respond to nutrients to regulate their metabolic and growth properties, which undoubtedly will be instrumental in the design of new therapeutic targets in cancer.